PROJECT SUMMARY The aim of this proposal is to characterize the clinical relevance of novel autoantibodies produced against neutrophil extracellular traps (NETs) in cystic fibrosis (CF). Lung complications are the main cause of mortality in cystic fibrosis (CF). CF lungs are characterized by chronic infiltration of neutrophils. Clinical data clearly show that released neutrophil-derived granule components and DNA are associated with lung function decline in CF. Results obtained by us and others indicate that formation of neutrophil extracellular traps (NETs) provides the major mechanism for the release of neutrophil-mediated inflammatory mediators in CF airways. Neutrophils release NETs to trap and kill extracellular microbes. However, uncontrolled NET release has been linked to several diseases such as CF. Besides having short-term damaging effects on surrounding tissues, released NET components could also serve as antigens triggering an autoimmune response in CF. Our preliminary data obtained in human CF clinical biospecimen using five independent approaches show that several autoantibodies produced against NET components are present in CF serum and sputum. This suggests that autoantibodies targeting NETs are produced in CF patients. Our long-term research goal is to characterize this novel autoimmune component of CF. The objective here, which is our logical first step in pursuit of that goal, is to establish that autoantibodies specific for NET components are present in CF and are associated with reduced lung function or other clinical symptoms. Our central hypothesis is that levels of anti-NET autoantibodies present in CF correlate with lung disease severity and impaired NET degradation of CF sera. To test our hypothesis, we will perform the following three specific aims: 1) Determine correlation of anti-NET autoantibodies with CF lung disease severity; 2) Determine whether CF patients have elevated levels of anti-citrullinated protein antibodies; and 3) Determine whether anti-NET autoantibodies lead to impaired NET degradation in CF sera. Our expected outcomes are that 1) CF lung disease severity correlates with serum levels of anti-NET antibodies and the inability of CF serum to degrade NETs; 2) anti-citrullinated antibodies characteristic for the autoimmune disease, rheumatoid arthritis, are also present in CF; 3) anti-NET autoantibodies are potentially linked to CF-related arthritis, vasculitis, kidney disease, diabetes or cutaneous manifestations. Overall, our work proposes that CF has an underappreciated autoimmune component likely induced by the chronic presence of NET-derived host molecules. Our proposed work will have a positive impact by providing essential, novel knowledge on the presence and clinical relevance of anti-NET autoantibodies in CF. It will advance our understanding of CF airway inflammation and autoimmunity in CF.